This proposal describes the framework of an R01 grant by Dr. Daniel Lingwood, an Assistant Professor at Harvard Medical School and faculty member of the Ragon Institute of MGH, MIT and Harvard, and Dr. Bryce Chackerian, an Associate Professor in the Department of Molecular Genetics & Microbiology at the University of New Mexico School of Medicine. Their research centers on B cell receptor (BCR) antigen recognition biology (Lingwood) and directed evolution of virus-like particle (VLP) vaccine platforms (Chackerian). Together they propose to develop a new strategy for universal influenza vaccine design. Most vaccine-elicited antibody responses to this virus are dominated by immunodominant off-target, non-neutralizing activities. However, recent work from Dr. Lingwood indicates that human BCRs assembled using the antibody gene IGHV1-69 possess V region-encoded (innate-like) specificity for a functionally conserved site of vulnerability, the stem- epitope of the influenza spike protein hemagglutinin (HA) and target of broadly neutralizing antibody (bnAb) responses. To experimentally evaluate this as a gene-encoded template for building a universal vaccine, Dr. Lingwood has generated transgenic mice in which antibody development proceeds via normal human VDJ recombination, but where V region use is constrained to IGHV1-69. Preliminary data indicate that IGHV1-69 usage itself refocuses the antibody response to the stem epitope, a feature that Dr. Lingwood finds is dependent on a single gene-encoded amino acid in the stem-contacting CDRH2 of IGHV1-69. To now transduce this into bnAb elicitation, Dr. Lingwood proposes to immunize his humanized mice with rationally- designed trimeric and nanoparticle displays of influenza HA stem which trigger innate-like stem-epitope signaling by the reconstituted germline IGHV1-69 BCR. Through these experiments Dr. Lingwood will provide a major paradigm shift in rational vaccine design, namely that broad protection may be generated through `activation' and `amplification' of gene-encoded antibody responses. To define a pathway for clinical development, Drs. Lingwood and Chackerian have applied RNA bacteriophage peptide display and affinity selection technology to derive a VLP vaccine with multivalent affinity to the V region of IGHV1-69 germline BCR, in essence a V region-specific primer to selectively expand innate-like stem targeting activity that is otherwise normally diluted by human BCR diversity. Selective IGHV1-69 priming will be evaluated in the Trianni mouse, the latest industry-standard humanized mouse vaccine model. V region priming will then be boosted with IGHV1-69-engaging HA stem immunogens, to stimulate a now immunodominant innate-like HA stem sensing antibody response. Exploiting a genetic basis for bnAb elicitation aims to overcome the failure of traditional approaches to influenza vaccination and is consistent with the purpose of this funding opportunity and broader mission of the NIH.